The Na.sup.+ /H.sup.+ exchangers, or antiporters, are plasma membrane transport proteins that exchange extracellular Na.sup.+ for intracellular H.sup.+ and are found in virtually all animals. In fact, all eukaryotic cells studied, including yeast, the worm (Caenorhabditis elegans), and crustaceans, have exhibited plasma membrane Na.sup.+ /H.sup.+ exchangers (also called NHE) which exchange these ions at a ratio of 1:1. Prokaryotes have functionally similar Na.sup.+ /H.sup.+ exchanger proteins which also regulate intracellular Na.sup.+ ion concentration and pH, and exchange one intracellular Na.sup.+ for one H.sup.+.
In eukaryotic cells, the plasma membrane Na.sup.+ /H.sup.+ exchangers have multiple functions, including pH homeostasis, volume regulation, cell proliferation, and transcellular Na.sup.+ absorption. In no cell, however, is the Na.sup.+ /H.sup.+ exchanger the only mechanism for these functions. For instance, pH homeostasis is controlled in most eukaryotic cells by mechanisms including a Cl.sup.- /HCO.sub.3.sup.- exchanger, a NaHCO.sub.3.sup.- co-transporter, a NA.sup.+ -dependent Cl.sup.- /HCO.sub.3.sup.- exchanger, and multiple mechanisms of H.sup.+ extrusion.
Nonetheless, understanding the Na.sup.+ /H.sup.+ exchanger will greatly increase the understanding of the body's control of ions, and much work has been done on the Na.sup.+ /H.sup.+ exchanger family. Four mammalian Na.sup.+ /H.sup.+ exchanger isoforms have been cloned (NHE1-4) (28). Of these, NHE3 appears to be the Na.sup.+ /H.sup.+ exchanger isoform that is most likely responsible for "brush border" Na.sup.+ /H.sup.+ exchange activity.
The brush border consists of microvilli, approximately 1.mu. in length and 0.1.mu. in diameter, that protrude from the surface of epithelial cells on the intestine and renal tubules. These microvilli greatly increase the surface area of those cells. Brush border Na.sup.+ /H.sup.+ exchange activity contributes to transepithelial neutral NaCl absorption (38) in the small intestine, and to Na.sup.+ reuptake in the proximal renal tubule (11,12,16). Additionally, brush border Na.sup.+ /H.sup.+ exchange activity is important in the secretion of acid in the proximal renal tubule (1).
Thus, a malfunctioning Na.sup.+ /H.sup.+ exchanger affects a body's well being. In chronic metabolic acidosis, chronic renal failure, diabetic nephropathy, and in animal models of essential hypertension, one observes an increase in renal proximal tubule brush border Na.sup.+ /H.sup.+ exchange activity (16,18). More specifically regarding hypertension, increased Na.sup.+ /H.sup.+ exchange in the renal proximal tubule or cortical thick ascending limb of Henle would enhance Na.sup.+ reabsorption, leading to a defect in renal Na.sup.+ excretion (16). It has been shown that defective renal Na.sup.+ excretion is a cause in some patients of essential hypertension (16).
Similarly, increased jejunal brush border Na.sup.+ /H.sup.+ exchange has been shown to be present in animal models of essential hypertension (40). Increased ileal and renal brush border Na.sup.+ /H.sup.+ exchange activity is an important mechanism for the increased ileal and renal NaCl and water absorption that occurs in response to administration of glucocorticoids (34,41) and thus may, in part, be responsible for common side-effects of glucocorticoid pharmacologic therapy in humans such as hypertension and fluid and salt retention.
Conversely, decreased brush border Na.sup.+ /H.sup.+ exchange activity is the major mechanism for decreased Na.sup.+ and water absorption in most human diarrheal diseases (21). In one familial diarrheal syndrome, congenital sodium diarrhea, there is evidence of a congenital absence of jejunal brush border Na.sup.+ /H.sup.+ exchange activity (4).
NHE3 is believed to be the Na.sup.+ /H.sup.+ exchanger that is increased in the above renal diseases and inhibited in diarrheal diseases because the Na.sup.+ /H.sup.+ exchange activity of NHE3 is most like that of ileal villus cell brush border membranes: it is relatively resistant to Na.sup.+ /H.sup.+ exchange inhibition by amiloride, and it is the only Na.sup.+ /H.sup.+ exchanger isoform inhibited and not stimulated by protein kinase C (28). Furthermore, only NHE3 message expression increases in parallel with the increased ileal villus apical Na.sup.+ /H.sup.+ exchange activity in rabbits treated for 24h with methylprednisolone (34). Recent immunohistochemical studies and Western analysis have demonstrated that NHE3 is present on the brush border but not the basolateral membranes of ileal villus and ascending colon surface epithelial cells and proximal renal tubules (2,3). Therefore, NHE3 is believed to be the Na.sup.+ /H.sup.+ exchanger isoform responsible for the characteristic Na.sup.+ /H.sup.+ exchange activity of the brush border membranes of the mammalian small intestinal, colonic and proximal renal tubule Na.sup.+ absorbing cells (14,28).
In addition, activation of plasma membrane Na.sup.+ /H.sup.+ exchange has been postulated to play an important role in oncogenic transformation, and inhibitors of plasma membrane Na.sup.+ /H.sup.+ exchange activity have been shown to have antitumoral effects (16,42).
A partial human NHE3 cDNA, clone HKC-3, which encodes 325 amino acids having 94% identity to rabbit NHE3 residues 180-505, has been previously reported (26). Clone HKC-3 has been used to physically and genetically map human NHE3 to chromosome 5p15.3, making NHE3 the most telomeric gene identified on chromosome 5p (6) and proving that the human NHE3 homologue arises from a different gene than human NHE1, mapped to 1p (15). Nonetheless, the existence of the clone did not provide the full DNA sequence of human NHE3 or the protein itself, and there remained in the art a need for that sequence.